The invention relates to DNA sequences for human matrix metalloproteases as well as to homologous sequences derived therefrom. It furthermore relates to the proteins and protein variants, coded by the DNA sequences, their expression, preparation and utilization. Areas of application are molecular biological, medical and pharmaceutical research, medical diagnosis and therapy and the pharmaceutical and biotechnological industry.
Matrix metalloproteases hydrolyze proteins of the extracellular matrix. They change the matrix structure and effect cell-matrix interactions. The matrix metalloproteases include collagenases, gelatinases, stromelysins and metalloelastases [1]. The following are some of the physiological processes, in which the enzymes participate: ovulation [2], embryo implantation in the uterus [3], cell migrations and tissue inversions during embryo genesis [4], involution of the mammary gland [5] and of the uterus [6] and angiogenesis [7]. Matrix metalloproteases play an important role in wound healing and scar formation [8], in metastasizing of tumors cells [9, 10], in rheumatic arthritis and osteoarthritis [11, 12] and in periodontal diseases [13].
All matrix metalloproteases contain a Zn.sup.2+ ion in the active center. The activation of the matrix metalloproteases, synthesized in the form of inactive proenzymes, requires the dissolution of a bond between the Zn.sup.2+ ion in the active center and a Cys group in the N-terminal propeptide of matrix metalloproteases (cysteine switch) [14]. Matrix metalloproteases consist of several protein domains, which exhibit homology among members of the protease family [1, 14]. Whereas the protease matrilysin consists only of a propeptide and of the amino acid sequence of the catalytic domain, other matrix metalloproteases contain, in addition, a hemopexin-like sequence of about 200 amino acids. The gelatinases A and B contain additional amino acid sequences. Known human matrix metalloproteases, their molecular weights and their preferred substrates are listed in Table 1.
TABLE 1 ______________________________________ MATRIX METALLOPROTEASES Protease M.sub.r (kDa) Substrate ______________________________________ Interstitial 54.1 Collagen I, II, III Collagenase (MMP-1) Neutrophilic 53.4 Collagen I, II, III Collagenase (MMP-5) Gelatinase A 73.9 Collagen IV, V, VII (MMP-2) Gelatin, Elastin Gelatinase B 78.4 Collagen IV, V (MMP-9) Gelatin, Elastin Stromelysin 1 54 Proteoglycans, (MMP-3) Fibronectin, Laminin, Gelatin, Collagen II, IV, V, IX Stromelysin 2 54.1 Proteoglycans, (MMP-10) Fibronectin, Laminin, Gelatin, Collagen II, IV, V, IX Matrilysin 29.7 Proteoglycans, (MMP-7) Fibronectin, Gelatin, Elastin Stromelysin 3 54.6 Metalloelastase 54 Fibronectin, Elastin ______________________________________
The different matrix metalloproteases are distinguished not only by a characteristic, macromolecular specificity for matrix proteins. Their activity is controlled on different molecular and cellular level:
1. Regulation of the synthesis of matrix metalloproteases by growth factors, cytokines, polypeptide hormones, prostaglandins, glucocorticoids, estrogen, progesterone and other effectors [1, 14]. PA1 2. Binding of matrix metalloproteases to membrane receptors [15]. PA1 3. Activation of inactive proenzymes by specific hydrolysis of the respective propeptides [16] or by oxidation [17]. PA1 4. Inhibition of matrix metalloproteases by specific protein inhibitors such as TIMP-I, TIMP-2 and TIMP-3 (TIMP=Tissue Inhibitor of Matrix Metalloproteases) [16]. PA1 5. Proteolytic degradation of matrix metalloproteases.
Matrix metalloproteases are being investigated intensively because of their important physiological functions and their role in the pathogenesis of diseases. There is interest in finding and characterizing further matrix metalloproteases.